The malignant B-lineage Reed-Sternberg (H-RS) cells in about half of all cases of Hodgkin lymphoma express Epstein-Barr virus (EBV) antigens, rendering this tumor highly attractive for cytotoxic T-lymphocyte (CTL) immunotherapy. However, these tumor-associated viral antigens are either subdominant (LMP1 and LMP2) or not presented to the immune system (EBNAI and BARFO). The H-RS cells also express molecules that are inhibitory to CTL, such as TGF-beta, the chemoattractant TARC and Fas-ligand that can comprise immunotherapy even when immune epitopes are effectively presented to CTLs. The studies proposed here seek to overcome these immune evasion tactics by redirecting immune responses to the LMP2 viral antigen, using LMP2a-transduced dendritic cells, and by genetically modifying CTLs in vitro so that they will resist the inhibitory effects of TGF-beta and Fas ligand. The latter aim will rely on a transgenic dominant-negative TGF-beta type 2 receptor expressed on CTLs and the adenoviral RID protein, which should render CTLs resistant to inhibition by TGF-beta and killing through Fas ligand-receptor interactions. Our preliminary studies suggest that there may be few T helper epitopes in LMP2. Since persistence in vivo is dependent on the availability of help, we will also explore the possibility that EBNA1, expressed as a retrogen in dendritic cells (using technology developed in project 4) will reactivate EBNA1-specific CD4+ T cells that can provide cognate help for LMP2-specific CTL as well as broaden the cytotoxic repertoire of the tumor specific CTL by reactivating CD4+ EBNA1-specific CTL. The hypotheses underlying these aims will be tested both in vitro and in vivo (Phase l/ll trials in patients with post-transplant or relapsed EBV-positive Hodgkin Lymphoma). Upon successful completion of the project, we will have learned whether our experimental modifications are safe in patients and whether they will render CTLs resistant to several of the immune evasion strategies commonly used by tumor cells.